Objective:To investigate the essential determinants that are critical to initiating the assembly of outer membrane proteins by replacing the POTRA domains of the translocator protein FhaC and the insertase protein TtOmp85 of the Omp85 family. Methods:FhaC, TtOmp85 proteins and their recombinant chimeric proteins after replacing the POTRA domain were obtained by overexpression and purification in vitro. An in vitro reconstitution system was used to investigate the effects of the different domains on the transport efficiency of the substrate outer membrane protein FhaB and the membrane insertion efficiency of OmpA. Results:Replacing the POTRA domain of FhaC with that of TtOmp85 led to the loss of the transport function of FhaB. During the membrane insertion process of OmpA, the FhaC mutant containing the POTRA of TtOmp85 protein acquired the ability to assemble OmpA. Conclusion:The compositional differences in the POTRA domain of Omp85 family proteins determine their abilities to recognize their substrate proteins.

Objective:To investigate the essential determinants that are critical to initiating the assembly of outer membrane proteins by replacing the POTRA domains of the translocator protein FhaC and the insertase protein TtOmp85 of the Omp85 family. Methods:FhaC, TtOmp85 proteins and their recombinant chimeric proteins after replacing the POTRA domain were obtained by overexpression and purification in vitro. An in vitro reconstitution system was used to investigate the effects of the different domains on the transport efficiency of the substrate outer membrane protein FhaB and the membrane insertion efficiency of OmpA. Results:Replacing the POTRA domain of FhaC with that of TtOmp85 led to the loss of the transport function of FhaB. During the membrane insertion process of OmpA, the FhaC mutant containing the POTRA of TtOmp85 protein acquired the ability to assemble OmpA. Conclusion:The compositional differences in the POTRA domain of Omp85 family proteins determine their abilities to recognize their substrate proteins.

Objective:To screen active antibacterial components from licorice extract using BamA and BamD, the core components of Escherichia coli ( E. coli) β-barrel assembly machinery (BAM), as targets in order to combat the increasingly serious problem of antibiotic resistance. Methods:Affinity ultrafiltration combined with high performance liquid chromatography-mass spectrometry (HPLC-MS) was used to screen the potential components interacting with BamA and BamD from licorice extract. Changes in the expression of bamA and bamD genes of E. coli after treatment with the compounds were detected by fluorescence quantitative PCR, and the effects of the compounds on the function of the BAM complex to integrate outer membrane proteins into the bacterial outer membrane were analyzed using an in vitro recombination system. The influence of the compounds on the integrity of bacterial membranes was evaluated through analyzing the accumulation of SDS within the bacterial cells. Results:Bioaffinity ultrafiltration combined with HPLC-MS screening revealed that 18β-glycyrrhetinic acid could interact with BamD. After 18β-glycyrrhetinic acid treatment, the expression of bamA gene increased by 1.5 times, and the expression of bamD gene increased by 2 times. However, the inhibitory effect of 18β-glycyrrhetinic acid on the membrane insertion function of the BAM complex was not observed in the in vitro recombinant system assay, and the cell membrane integrity assay experiments did not reveal any disruption of the E. coli cell membrane by 18β-glycyrrhetinic acid. Conclusions:Using BamA and BamD proteins as targets, a natural product screening method using affinity ultrafiltration combined with HPLC-MS is established. The screening result shows that 18β-glycyrrhetinic acid can interact with BamD and affect the expression of outer membrane proteins in E. coli. Therefore, the screening and experimental procedures established in this study are of good reference value for the screening of novel antimicrobial drugs from other sources targeting outer membrane proteins, and this study also suggests that the selection of the relevant target sites is crucial for the successful screening of the corresponding natural products.

Objective:To investigate whether there is a correlation between right bundle branch block (RBBB) and nonalcoholic fatty liver disease (NAFLD) in patients with type 2 diabetes mellitus.Methods:A retrospective analysis of 226 patients with type 2 diabetes mellitus who were admitted to the Fifth Affiliated Hospital of Zhengzhou University from March 2018 to July 2019 was performed. According to the results of electrocardiogram examination, they were divided into RBBB group ( n=58 patients) and non-RBBB group ( n=168 patients). The general clinical data of the two groups of patients were collected, blood lipids, liver function, renal function, coagulation function and other related indicators were measured on the fasting of the next morning. The diagnosis of NAFLD is based on ultrasound. Logistic regression analysis was performed on factors that may affect RBBB. Results:Of the 226 patients with type 2 diabetes mellitus, 127 (56.2%) were male and 99 (43.8%) were female. The composition of male, age, diabetes duration, hypertension, fibrinogen (FIB), serum creatinine (SCr), alanine aminotransferase (ALT), and NAFLD in the RBBB group was higher than that in the non-RBBB group ( P<0.05). The levels of total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C) and albumin (ALB) in the RBBB group were lower than those in the non-RBBB group ( P<0.05). Logistic regression analysis showed that male ( OR=2.736, 95% CI: 1.075-5.251, P=0.032), advanced age ( OR=1.049, 95% CI: 1.009-1.090, P=0.016), higher serum creatinine levels ( OR=1.045, 95% CI: 1.021-1.070, P<0.001), and NAFLD ( OR=2.834, 95% CI: 1.166-6.891, P=0.022) were independent risk factors of RBBB in patients with type 2 diabetes mellitus. Conclusions:NAFLD may be associated with an increased risk of right bundle branch block in patients with type 2 diabetes.

Objective To investigate the effect of recombinant human endostatin combined with chemotherapy on the expression of vascular endothelial growth factor (VEGF) in patients with advanced cancer.Methods250 patients with advanced malignant tumors were randomly selected from February 2012 to February 2016 the Third Hospital of Jiaxing, with unit control method experiment principle divided into two groups: experimental group (125 cases) and control group (125 cases).The experimental group treated by recombinant human endostatin combined chemotherapy, control group received routine chemotherapy for a week during the period of 21 days, after two cycles of treatment, clinical efficacy and adverse reactions of the two groups of patients were compared, and the expression of VEGF in tumor blood vessels weredetected.ResultsAfter two cycles of treatment, objective response rate, disease control rate in the experimental group patients was significantly higher than that in control group (P<0.05);after treatment, the expression level of VEGF in serum of the experimental group and the control groupwere decreased significantly, but the experimental group was lower than that of control group was more obvious (P<0.05).ConclusionRecombinant human endostatin combined with chemotherapy in the treatment of advanced malignant tumors can enhance the effect of chemotherapy, regulate the level of VEGF.